Coordinated in vitro and in vivo studies of biochemical and pharmacokinetic aspects of folate analog pharmacology are proposed in murine tumor models. It continues our earlier studies which provided evidence for a role of membrane transport of folate analogs in their selective antitumor action. Studies initiated with L1210 cells will now consider questions as to differences in the multiplicity and structural specificity of systems transporting folate analogs in normal proliferative tissue and tumors differing in responsiveness. Specificity for diastereoisomers of reduced folates, homologs of 4-amino folates and folic acid and metabolites will be evaluated along with dihydrofolate reductase binding specificity. Quantitative aspects of the dynamics and interrelationship between the pharmacokinetics and membrane transport of folate analogs will be sought in different tissues during therapy. Transport parameters will be assessed by measuring intracellular accumulation in vivo, after removing cells from drug-treated mice and also by probing the kinetic interrelationships with pharmacologic agents such as probenacid. The extent and time-course for perturbations of dihydrofolate reductase level, of tetrahydrofolate synthesis and utilization and of dihydrofolate accumulation in various tissues will be explored in vivo during and after therapy and evaluated in respect to requirements for intracellular exchangeable levels of drug. Studies of polyglutamate formation in vivo from methotrexate will be continued and initiated with aminopterin to fully assess this metabolism in terms of the observed pharmacokinetics in each tissue. More rigorous assessment of the membrane permeability and hydrolysis of intracellular polyglutamates vis a vis unchanged analog will also be made. Biochemical studies will also be initiated in L1210 cells in culture seeking a basis of the schedule-dependent synergism between methotrexate and vincristine. Similar studies of the interaction of 5-FU with the 5-arylpyrimidine antifolate, DDMP, will also be initiated to ascertain synergism similar to that demonstrated to 5-FU with methotrexate.